The contribution of enzyme adaptation and substrate availability to the regulation of urea synthesis will be studied. Isolated rat hepatocytes will be used to determine the subcellular distribution of urea cycle intermediates; different fluxes through the urea cycle will be generated in order to establish whether that distribution changes under different conditions, and to observe the patterns of change in the liver content of urea cycle and related intermediates at different ammonia loads. This information will be used to analyze the data obtaine from the following proposed work. The activity of the urea cycle enzymes in the liver and kidneys of rats fed isocaloric diets containing different proportions of protein will be measured, as well as the liver content of urea cycle intermediates and related metabolites in similarly treated animals. In addition, for each nutritional condition, the effect of a sudden increase in the ammonia load on the level of the relevant intermediates in liver will be determined. Analysis of the data obtained will provide information on some of the mechanisms which contribute to the short- and long-term regulation of urea synthesis in the liver, and on the possible role the kidney may play in the sparing of arginine under conditions of increased degradation of endogenous protein.